Expression of recombinant proteins in microbial host cells is widely used for industrial production of human therapeutic proteins. A fusion partner may be attached to the N-terminal (or C-terminal) of the target protein and serves to increase expression level, solubility or correct folding of the target protein and to facilitate easy purification during downstream processing. Removal of the fusion partner from the target protein is mostly necessary in order to conserve the biological activity of the target protein. The processing of the fusion protein can either be performed after expression or after one or more initial purification steps using suitable proteases capable of cleaving the fusion partner to release the target protein. The recombinant target protein with or without a fusion partner may also need to be modified after translation to obtain certain biological features using one or more enzymatic steps. Such modification includes removal of unwanted glycosylations with mannose, fucose or xylose sugars or introductions of glycosylations. Modifications may also include deamidations, amidations, methylations, phosphorylations, dephosphorylations, sulfations, acetylations or transaminations.
To decrease the cost of the down stream processing steps it is pivotal that these processing enzymes are cheap to produce e.g. can be expressed in high amounts and purified with a small number of chromatographic steps. Furthermore it is desirable that processing enzymes used for cleavage of fusion proteins can be easily removed after processing of the target protein. It may also be an advantage that the processing enzymes can be immobilised to purifications columns.
The present invention provides novel processing enzymes which are well suited for modification of target proteins including cleavage of fusion partners from a fusion protein. Furthermore, the novel processing enzymes can easily be separated from the target protein after the processing step.